Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.4.1.2 (glutamate dehydrogenase)
4,380 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effect of chronic acid feeding and its subsequent withdrawal was determined on the amounts of the metabolic intermediates and enzymic activities of the purine nucleotide cycle. Sprague-Dawley rats were given 1.5% (w/v) NH4Cl in their drinking water for 5 days. The renal excretion of NH3 rose 70-fold and the rats developed acidosis. The amount of renal IMP rose from a control value of 4.5 +/- 2.2 to 20.4 +/- 3.7nmol/g of kidney after 48h of acid feeding (P less than 0.001) and fell to normal within 48h of the recovery. Adenylosuccinate concentrations fell from a control value of 4.5 +/- 0.9nmol/g of kidney to 1.2 +/- 0.3nmol/g (P less than 0.005) by day 5 of acidosis and continued to fall to undetectable values by 48h after recovery. The amount of AMP remained constant through the acid-feeding and the recovery periods. The activity of adenylosuccinate synthetase, the rate-limiting enzyme of the purine nucleotide cycle, paralleled the rise and fall in NH3 excretion. The activities of phosphate-dependent glutaminase and glutamate dehydrogenase were elevated during the acid-feeding and the recovery period. Thus changes in the purine nucleotide cycle correlate with changes in NH3 excretion to a more parallel degree than does the activity of glutaminase or glutamate dehydrogenase.
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PMID:The purine nucleotide cycle in the regulation of ammoniagenesis during induction and cessation of chronic acidosis in the rat kidney. 730 74

The effect of different adenine-containing compounds on the NADP-/NAD-glutamate dehydrogenase (GDH) ratio was studied as a function of yeast-mycelium transition in Benjaminiella poitrasii. Under in vivo conditions, at a 5.0 mM concentration, cyclic AMP (cAMP) and dibutyryl cAMP maintained the cells in the yeast form for up to 7 and 5 h, respectively, and this was reflected in the patterns of GDH ratios observed. In vitro studies of phosphorylation and dephosphorylation have also been carried out, and the results suggest a possible correlation between cAMP, the GDH ratio, and cell form in B. poitrasii.
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PMID:Possible involvement of cyclic adenosine 3',5'-monophosphate in the regulation of NADP-/NAD-glutamate dehydrogenase ratio and in yeast-mycelium transition of Benjaminiella poitrasii. 839 89

Photoaffinity labeling with [alpha-32P]8N3GTP and [gamma-32P]8N3GTP was used to identify the guanine binding domain of the GTP regulatory site within glutamate dehydrogenase (GDH). Without photolysis, 8N3GTP mimicked the regulatory properties of GTP on GDH activity with 8N3GTP exhibiting a Ki of 5 microM while the Ki for GTP was about 0.6 microM. Under optimal photolabeling conditions saturation of photoinsertion with 1 microgram of GDH revealed an apparent Kd of 9 +/- 4 microM for [gamma-32P]8N3GTP. Photolabeling with this analog could be competitively inhibited with GTP with an apparent Kd of 12 +/- 2 microM. Other nucleotides such as ATP and NAD(P)H could not reduce the amount of photoinsertion as effectively as GTP. ADP could decrease photoinsertion, but only at much higher concentrations. NAD(P)+, GDP, AMP, and GMP had little effect on photoinsertion. Divalent cations Mg2+ and Ca2+ also reduced photoinsertion significantly while the monovalent K+ and Na+ ions had no effect. Aluminum(III)-chelate or iron(III)-chelate affinity chromatography and reversed-phase HPLC were used to purify photolabel-containing peptides generated with either trypsin or chymotrypsin. This identified a portion of the guanine binding domain within the GTP regulatory site as the region containing the sequence Ile439 to Tyr454. Photolabeling of this peptide was prevented 91% by the presence of 300 microM GTP during photolysis. Lys445 was not identified in sequence analyses of the photolabeled peptides. Also, trypsin was unable to cleave the photolabeled peptide at this site. These results suggest that Lys445 may be the residue modified by [alpha-32P]8N3GTP.
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PMID:Identification of a guanine binding domain peptide of the GTP binding site of glutamate dehydrogenase: isolation with metal-chelate affinity chromatography. 843 45

A variety of metabolites have been found to elicit a form of inhibition or activation on an NAD-specific glutamate dehydrogenase (NAD-GDH, EC 1.4.1.2) from Halobacterium halobium. The purified halophilic enzyme was tested with several compounds known to be allosteric modifiers of mammalian glutamate dehydrogenases to determine their effects on enzyme activity. GTP, ATP, ADP and AMP did not affect the enzyme, so these effectors of bovine glutamate dehydrogenase do not play a role in the regulation of the halophilic enzyme. However, the halophilic enzyme was subject to strong inhibition by TCA intermediates. When measuring the initial rate of the reaction, the oxidative deamination of L-glutamate was inhibited by TCA metabolites such as: fumarate, oxalacetate, succinate and malate; by substrate analogues such as: NADP+, D-glutamate and glutarate; and by dicarboxylic compounds such as adipate. On the other hand, all the amino acids tested were activators of this enzyme, except the D-isomer of the substrate L-glutamate that acted as an inhibitor. The relative effectiveness of each inhibitor or activator (Ki or Ka values) was correlated with the dipole moment (mu), HOMO and LUMO molecular orbital energies, optimal distance between two carboxyl groups, and hydrophobicity. Compounds with high dipole moment acted as good activators while compounds with low dipole moment were inhibitors. We have also found that the best activators were amino acids with no polar lateral chain.
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PMID:NAD-glutamate dehydrogenase from Halobacterium halobium: inhibition and activation by TCA intermediates and amino acids. 860 24

Two forms of the NAD-dependent glutamate dehydrogenase were partially purified from Dictyostelium discoideum, an activated and a non-activated form. V(max) for the non-activated enzyme was stimulated 88-fold and the activated enzyme 3-fold by 0.1 mM AMP (at their pH optima). Half maximal stimulation by AMP is achieved at 221 +/- 39 microM for the non-activated enzyme and 20 +/- 2 microM for the activated enzyme. We have shown that activation of NAD-GDH in vivo has many similarities to trypsin treatment of non-activated enzyme and that proteolysis is the probable mechanism of activation.
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PMID:Kinetic properties and the mechanism of activation of NAD-dependent glutamate dehydrogenase from Dictyostelium discoideum. 872 2

The NAD-dependent glutamate dehydrogenase (GDH) (EC 1.4.1.2) from Laccaria bicolor was purified 410-fold to apparent electrophoretic homogeneity with a 40% recovery through a three-step procedure involving ammonium sulfate precipitation, anion-exchange chromatography on DEAE-Trisacryl, and gel filtration. The molecular weight of the native enzyme determined by gel filtration was 470 kDa, whereas sodium dodecyl sulfate-polyacrylamide gel electrophoresis gave rise to a single band of 116 kDa, suggesting that the enzyme is composed of four identical subunits. The enzyme was specific for NAD(H). The pH optima were 7.4 and 8.8 for the amination and deamination reactions, respectively. The enzyme was found to be highly unstable, with virtually no activity after 20 days at -75 degrees C, 4 days at 4 degrees C, and 1 h at 50 degrees C. The addition of ammonium sulfate improved greatly the stability of the enzyme and full activity was still observed after several months at -75 degrees C. NAD-GDH activity was stimulated by Ca2+ and Mg2+ but strongly inhibited by Cu2+ and slightly by the nucleotides AMP, ADP, and ATP. The Michaelis constants for NAD, NADH, 2-oxoglutarate, and ammonium were 282 &mgr;M, 89 &mgr;M, 1.35 mM, and 37 mM, respectively. The enzyme had a negative cooperativity for glutamate (Hill number of 0.3), and its Km value increased from 0.24 to 3.6 mM when the glutamate concentration exceeded 1 mM. These affinity constants of the substrates, compared with those of the NADP-GDH of the fungus, suggest that the NAD-GDH is mainly involved in the catabolism of glutamate, while the NADP-GDH is involved in the catalysis of this amino acid. Copyright 1997 Academic Press. Copyright 1997 Academic Press
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PMID:Purification and characterization of the NAD-dependent glutamate dehydrogenase in the ectomycorrhizal fungus laccaria bicolor (Maire) orton 945 44

A new class of glutamate dehydrogenase (GDH) is reported. The GDH of Streptomyces clavuligerus was purified to homogeneity and characterized. It has a native molecular mass of 1,100 kDa and exists as an alpha(6) oligomeric structure composed of 183-kDa subunits. GDH, which requires AMP as an essential activator, shows a maximal rate of catalysis in 100 mm phosphate buffer, pH 7.0, at 30 degrees C. Under these conditions, GDH displayed hyperbolic behavior toward ammonia (K(m), 33 mm) and sigmoidal responses to changes in alpha-ketoglutarate (S(0.5) 1.3 mm; n(H) 1.50) and NADH (S(0.5) 20 microm; n(H) 1.52) concentrations. Aspartate and asparagine were found to be allosteric activators. This enzyme is inhibited by an excess of NADH or NH(4)(+), by some tricarboxylic acid cycle intermediates and by ATP. This GDH seems to be a catabolic enzyme as indicated by the following: (i) it is NAD-specific; (ii) it shows a high value of K(m) for ammonia; and (iii) when S. clavuligerus was cultured in minimal medium containing glutamate as the sole source of carbon and nitrogen, a 5-fold increase in specific activity of GDH was detected compared with cultures provided with glycerol and ammonia. GDH has 1,651 amino acids, and it is encoded by a DNA fragment of 4,953 base pairs (gdh gene). It shows strong sequence similarity to proteins encoded by unidentified open reading frames present in the genomes of species belonging to the genera Mycobacterium, Rickettsia, Pseudomonas, Vibrio, Shewanella, and Caulobacter, suggesting that it has a broad distribution. The GDH of S. clavuligerus is the first member of a class of GDHs included in a subfamily of GDHs (large GDHs) whose catalytic requirements and evolutionary implications are described and discussed.
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PMID:A new class of glutamate dehydrogenases (GDH). Biochemical and genetic characterization of the first member, the AMP-requiring NAD-specific GDH of Streptomyces clavuligerus. 1092 16

Reactive oxygen species generated by xanthine oxidase during reperfusion of ischemic liver might in part be responsible for ischemic organ injury. In normothermic ischemia/reperfusion rat model, we investigated whether allopurinol pretreatment improved ischemia-induced mitochondrial dysfunction. Rats were subjected to 60 min of hepatic ischemia and to 1 h and 5 h of reperfusion thereafter. At 18 h and 1 h before ischemia, the animals received 0.25 mL of either saline or allopurinol (50 mg/kg) i.p. In saline-treated ischemic rats, serum aspartate aminotransferase levels increased significantly at 5 h (4685 +/- 310 IU/L) and were significantly reduced with allopurinol pretreatment. Similarly, mitochondrial lipid peroxidation was elevated in the saline-treated ischemic group, but this elevation was prevented by allopurinol. In contrast, mitochondrial glutamate dehydrogenase activity and ketone body ratio decreased in the saline-treated group, but this decrease was also inhibited by allopurinol. Hepatic ATP levels in the saline-treated rats were 42% lower 5 h after reperfusion. However, treatment with allopurinol resulted in significantly higher ATP levels. Allopurinol treatment preserved the concentration of AMP in ischemic liver but inhibited the accumulation of xanthine in reperfused liver. Our findings suggest allopurinol protects against mitochondrial injury, which prevents a mitochondrial oxidant stress and lipid peroxidation and preserves the hepatic energy metabolism.
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PMID:Protective effect of allopurinol on hepatic energy metabolism in ischemic and reperfused rat liver. 1122 Jun 38

The AMP-activated protein kinase (AMPK) exists as a heterotrimetric complex comprising a catalytic alpha subunit and non-catalytic beta and gamma subunits. Under conditions of hypoxia, exercise, ischemia, heat shock, and low glucose, AMPK is activated allosterically by rising cellular AMP and by phosphorylation of the catalytic alpha subunit. The mammalian target of rapamycin (mTOR) controls cellular functions in response to amino acids and growth factors. Recent reports including our study have demonstrated the possible interplay between mTOR and AMPK signaling pathways, supporting a model in which mitochondrial dysfunction caused by the mitochondrial inhibitors or ATP depletion inhibits activation of p70 S6 kinase alpha (p70alpha), a downstream effector of mTOR, by activating AMPK. Leucine may stimulate p70alpha phosphorylation via mTOR pathway, in part, by serving both as a mitochondrial fuel through oxidative carboxylation and an allosteric activation of glutamate dehydrogenase. This hypothesis may support an idea in which leucine modulates mTOR function, in part by regulating mitochondrial function and AMPK. Further understanding of the role of mTOR in coordinating amino acid- and energy-sensing pathways would provide new insights into relationship between nutrients and cellular functions.
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PMID:mTOR integrates amino acid- and energy-sensing pathways. 1468 82

The orphan receptor GPR80 (also called GPR99) was recently reported to be the P2Y(15) receptor activated by AMP and adenosine and coupled to increases in cyclic AMP accumulation and intracellular Ca(2+) mobilization (Inbe et al. J Biol Chem 2004; 279: 19790-9). However, the cell line (HEK293) used to carry out those studies endogenously expresses A(2A) and A(2B) adenosine receptors as well as multiple P2Y receptors, which complicates the analysis of a potential P2Y receptor. To determine unambiguously whether GPR80 is a P2Y receptor subtype, HA-tagged GPR80 was either stably expressed in CHO cells or transiently expressed in COS-7 and HEK293 cells, and cell surface expression was verified by radioimmunoassay (RIA). COS-7 cells overexpressing GPR80 showed a consistent twofold increase in basal inositol phosphate accumulation. However, neither adenosine nor AMP was capable of promoting accumulation of either cyclic AMP or inositol phosphates in any of the three GPR80-expressing cells. A recent paper (He et al. Nature 2004; 429: 188-93) reported that GPR80 is a Gq-coupled receptor activated by the citric acid cycle intermediate, alpha-ketoglutarate. Consistent with this report, alpha-ketoglutarate promoted inositol phosphate accumulation in CHO and HEK293 cells expressing GPR80, and pretreatment of GPR80-expressing COS-7 cells with glutamate dehydrogenase, which converts alpha-ketoglutarate to glutamate, decreased basal levels of inositol phosphates. Taken together, these data demonstrate that GPR80 is not activated by adenosine, AMP or other nucleotides, but instead is activated by alpha-ketoglutarate. Therefore, GPR80 is not a new member of the P2Y receptor family.
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PMID:GPR80/99, proposed to be the P2Y(15) receptor activated by adenosine and AMP, is not a P2Y receptor. 1840 2


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